人结肠癌羟基喜树碱多药耐药细胞的蛋白质组学研究

Proteomic study of hydroxycamptothecin-resistant human colon cancer cell line

目的对结肠癌羟基喜树碱多药耐药细胞(SW1116/HCPT)及其亲代细胞(SW1116)进行蛋白质组学比较研究.探讨肿瘤细胞的多药耐药机制。方法培养羟基喜树碱多药耐药细胞和亲代细胞.提取蛋白质.以固相pH梯度等电聚焦为第一向.十二烷基磺酸钠-聚丙烯酰胺凝胶垂直电泳为第二向进行双向电泳.图像分析软件分析电泳图谱,基质辅助激光解吸电离飞行时间质谱或基质辅助激光解吸电离飞行时间/飞行时间串联质谱鉴定蛋白质。结果发现9个蛋白质点在SW1116/HCPT细胞中表达量发生改变,4个蛋白质点表达量增加,5个蛋白质点表达量减少。质谱鉴定了6个蛋白质点分别为:琥珀酸脱氢酶复合物(亚单位A)、3-磷酸甘油醛脱氢酶、泛素融合降解1相似蛋白、胞核氯离子通道蛋白、β-微管蛋白和ORF蛋白。结论该研究有助于深入理解结肠癌羟基喜树碱多药耐药机制,并有望在新型耐药逆转剂的开发上发挥作用。

Objective To compare the results of proteomics between hydroxycamptothecin-resistant human colon cancer cells and their parent cells and to investigate multi-drug resistance mechanisms of digestive tract tumor. Methods Proteins were isolated from SW1116/HCPT cells and SW1116 cells and separated by two-dimensional（2D） gel electrophoresis. The proteins were then stained with silver or colloidal coomassie blue to produce a high-resolution map of the proteome. Selected proteins from this map were in-gel digested with trypsin and the resulting tryptic peptides were analyzed by Matrix assisted laser desorption-ionization（MALDI） time-of-flight（TOF） mass spectrometry and MALDI-TOF/ TOF tandem mass spectrometry. The mass spectrometric data was used to identify the proteins through search of the SWlSSPROT protein sequence database. Results There were nine proteins whose expres- sion level were changed in SWll16/HCPT cells as compared with SWl116 cells. Four were up-regulated and five were down-regulated. These protein spots were identified as succinate dehydrogenase complex（subunit A）, glyceraldehyde-3-phosphate dehydrogenase, ubiquitin fusion-degradation 1 like protein, nuclear chloride channel protein, tubulin beta and ORF protein. Conclusions There are differential expression of many drug resistant proteins in SWl116/HCPT cells. A number of related-proteins of drug resistance was obtained. Some proteins are quite useful for discovering the molecular mechanisms of multi-drug resistance.